Organization of the biosynthetic gene cluster for the polyketide anthelmintic macrolide avermectin in Streptomyces avermitilis

Abstract

Analysis of the gene cluster from Streptomyces avermitilis that governs the biosynthesis of the polyketide anthelmintic avermectin revealed that it contains four large ORFs encoding giant multifunctional polypeptides of the avermectin polyketide synthase (AVES 1, AVES 2, AVES 3, and AVES 4). These clustered polyketide synthase genes responsible for avermectin biosynthesis together encode 12 homologous sets of enzyme activities (modules), each catalyzing a specific round of polyketide chain elongation. The clustered genes encoding polyketide synthase are organized as two sets of six modular repeats, aveA1-aveA2 and aveA3-aveA4, which are convergently transcribed. The total of 55 constituent active sites makes this the most complex multifunctional enzyme system identified to date. The sequenced DNA region contains 14 additional ORFs, some of which encode polypeptides governing other key steps in avermectin biosynthesis. Between the two sets of polyketide synthase genes lie two genes involved in postpolyketide modification, one of which encodes cynthochrome P450 hydroxylase that probably catalyzes furan ring formation at C6 to C8a. Immediately right of the large polyketide synthase genes is a set of genes involved in oleandrose biosynthesis and its transglycosylation to polyketidederived aglycons. This cluster includes nine genes, but one is not functional in the biosynthesis of avermectin. On the left side of polyketide synthase genes, two ORFs encoding methyltransferase and nonpolyketide synthase ketoreductase involved in postpolyketide modification are located to the left of the polyketide synthase genes, and an adjacent gene encodes a regulatory function that may be involved in activation of the transcription of avermectin biosynthetic genes.Streptomyces avermitilis produces avermectin, a series of eight related pentacyclic lactones that contain a disaccharide of the methylated deoxysugar oleandrose (1). Avermectin and the related compounds milbemycin and nemadectin are potent anthelmintic compounds, the former two of which are used commercially in animal healthcare and agriculture. The semisynthetic derivatives of avermectin C22, C23 dihydroavermectin B1, ivermectin, are widely used for the treatment of diseases caused by nematodes and arthropods in veterinary and agricultural fields, respectively. Ivermectin has been used for livestock farming and health care of companion animals.

Characterization of bafilomycin biosynthesis in Kitasatospora setae KM-6054 and comparative analysis of gene clusters in Actinomycetales microorganisms

“…This loading AT-ACP didomain was identified at the N terminus of BfmA1, adjacent to the KS domain of module 1. A similar loading AT-ACP didomain has been reported in erythromycin 23 and avermectin 24 PKSs. Thus, all KS domains in bafilomycin PKS modules were functional (Supplementary Figure S1).…”

The autoregulator receptor homologue AvaR3 plays a regulatory role in antibiotic production, mycelial aggregation and colony development of Streptomyces avermitilis

“…Because the biosynthetic pathways of both avermectin and filipin require malonyl-CoA and methylmalonyl-CoA as a common extender unit of the polyketide backbone (Ikeda et al, 1999;Omura et al, 2001), it is most likely that the delayed expression of the ave genes, attributed to the delayed and weakened transcription of aveR as the result of avaR3 disruption, would lead to the preferential use of precursors in filipin production at the middle growth stage, resulting in increased filipin production and a lack of the necessary precursors for avermectin production at the late growth stage. Regarding oligomycin production, although oligomycin biosynthesis shares common precursors (malonyl-CoA and methylmalonyl-CoA) with avermectin and filipin biosynthesis (Wei et al, 2006), because oligomycin production already reached a plateau before the onset of avermectin production in the wild-type strain ( Supplementary Fig.…”

“…The biosynthetic pathways of the three types of antibiotic have been predicted and partially elucidated (Ikeda et al, 1999;Omura et al, 2001), while genetic information on the regulatory mechanism of antibiotic production has so far been limited. With respect to avermectin production, we recently reported that AveR, which is encoded by a gene situated on the left-hand extremity of the avermectin biosynthetic gene cluster, is a positive LAL-family regulator for controlling avermectin biosynthetic genes .…”

Insights into polyketide biosynthesis gained from repurposing antibiotic-producing polyketide synthases to produce fuels and chemicals

“…Branched-chain starter substrates in the polyketide chain initiation reaction are required for production of highly branched polyketides. It was known that the acyltransferase (AT)-acyl carrier protein (ACP) loading didomain of the avermectin PKS could use a variety of molecules, including the branched chain acyl-CoAs, isobutyryl-and 2-methylbutyryl-CoA, 6,7 to initiate avermectin biosynthesis, but we were not able to produce a soluble, active loading didomain either as a stand-alone protein or linked to an extender module (module 1) that would give us the required diketide in vitro. Hence we turned to the lipomycin PKS, which was proposed to use isobutyryl-CoA to initiate α-lipomycin biosynthesis, and whose AT-ACP loading didomain is linked to module 1 to form the first polypeptide (LipPks1) of the PKS complex ( Figure 1a).…”